Protective Effect And Mechanism Of HSYA On Coronary Artery Endothelial Cell Injury

Objective:The aim of this study was to examine the protective effect of the Guhong injection and its effective active ingredient hydroxysafflor yellow A?HSYA?on human coronary artery endothelial cells?HCAEC?induced by hydrogen peroxide?H₂O₂?and high oxidized low density lipoprotein?ox-LDL?.Methods:We simulated the process of atherosclerosis by treating HCAEC with high ox-LDL induction.Firstly,we examined the effects of different concentrations of Guhong injection,H₂O₂,ox-LDL,HSYA and high ox-LDL on the activity of HCAEC by methyltetrazolium blue?MTT?colorimetry,determined the optimal concentration of each drug,and establish the cell injury model.Secondly,we used nitrate reduction method and lactate dehydrogenase colorimetric method to detect the release of nitric oxide?NO?and dehydrogenated plum?LDH?in cell culture medium after treatment with high ox-LDL and HYSA.Next,TUNEL?TdT-mediated DUTP Nick-End Labeling?method was used to observe the apoptosis of different treatment groups.Finally,Western blotting?WB?and real-time reverse transcription polymerase chain reaction?RT-qPCR?were used to detect the expression of lectin-like oxidized low density lipoprotein receptor-1?LOX-1?and endothelial nitric oxide synthase?eNOS?proteins and mRNA in different treatment cell groups.Results:Compared with the control group,different concentrations of H₂O₂ acted on HCAEC,the cell survival rate decreased with the increase of concentration,and the cell survival rate increased after the intervention of Guhong injection,the difference was statistically significant?P<0.05,n=15?.Compared with the control group,with the increase of ox-LDL and high ox-LDL concentration,the cell survival rate decreased gradually.After co-treatment with Guhong injection or HSYA,the cell survival rate increased significantly,and the difference was statistically significant?P<0.01,n=15?.Compared with the control group,the cytosolic NO content in the high ox-LDL group decreased,and the LDH release increased significantly.Compared with the high ox-LDL group,the NO release in the high ox-LDL+HSYA group increased,and the LDH release decreased.All were statistically significant?P<0.01,n=9?.The TUNEL technique showed that the number of apoptotic cells in the high ox-LDL group was significantly increased.The number of apoptosis in the high ox-LDL+HSYA group was significantly decreased,and the apoptosis rate decreased from 46.16%to 31.88%.The difference was statistically significant?P<0.01,n=6?.Compared with the control group,the expression of eNOS protein was decreased and the expression of LOX-1 protein was up-regulated in the high ox-LDL group.At the same time,compared with the high ox-LDL group,the expression of eNOS protein was up-regulated and the expression of LOX-1 protein was down-regulated in the high ox-LDL+HSYA group,the difference was statistically significant?P<0.01,n=3?.Compared with the control group,the expression of eNOS mRNA was decreased and the expression of LOX-1 mRNA was up-regulated in the high ox-LDL group.At the same time,compared with the high ox-LDL group,the expression of eNOS mRNA was up-regulated and the expression of LOX-1 mRNA was down-regulated in the high ox-LDL+HSYA group,the difference was statistically significant?P<0.01,n=9?.Conclusion:Guhong injection can inhibit HCAEC injury induced by H₂O₂ and high ox-LDL.The HSYA inhibited the high ox-LDL-induced HCAEC injury and protected and promoted the cell repaired,possibly by up-regulating the eNOS gene and protein expression,increasing NO release,inhibiting LDH release and down-regulating LOX-1 mRNA and protein expression to achieve the effect.